Mycology Update 2025 - Margie Morgan, PhD

Mycology Update 2025
Margie Morgan, PhD, D(ABMM)
Mucormycetes Conidial Molds
Dematiacious Hyaline
Yeast
1

Starting points
Yeast:
• Unicellular and bud to produce daughter cells
• Growth on solid media usually produce white to beige colonies in the first
24-48 hours (30*-35*C) and appear much like bacterial colonies
• A few genera produce pigment (ex. Rhodotorula = orange/red)
• Some genera produce mucoid colonies (ex. Cryptococcus)
• Taxonomic updates: new name (old name) in parentheses)
Candida albicans
(5% Sheep’s blood agar)
Growth at 24hr / 35*C
2
C. neoformans
mucoid colonies
(Sabouraud agar)

Starting points
Molds:
• Produce hyphae and conidia [spores]
• Growth on solid media produce downy, fluffy, or cottony colonies
• Growth time is variable (3 – 21 days at 30-35*C) depending on genera of mold
• Mold colonies usually pigmented, which can assist with identification
• Taxonomic updates: new name (old name) in parentheses).
hyphae
spores
3

Specimen Collection and Transport
• Fungi are hardy
• Transport media usually not required
• Sterile containers used to prevent bacterial contamination /
room temperature transport is adequate
• Numerous anatomic sites are appropriate for culture:
• Respiratory specimens – sputum, bronchial lavage, brushings, nasal
sinuses
• Tissue biopsies
• Cutaneous - skin scrapings, material from lesions
• Ocular
• Sterile body fluids, including CSF
• Blood
• Bone Marrow
4

Fungal Culture Media / most common
• Sabouraud dextrose agar (SABS)
All purpose fungal medium – antibiotic free
Will also grow bacteria from contaminated sites
Contains 2% glucose, pH @7.
Best used for the subculture of fungi
for identification testing and susceptibility preparation
• Inhibitory mold agar (IMA)
Selective and enriched medium - contains antibiotics
chloramphenicol and gentamicin to inhibit bacterial growth
Good medium for the primary recovery of molds
5

Fungal Culture Media
• Mycosel (Mycobiotic) agar
• Highly selective SABs-like medium containing chloramphenicol and cycloheximide
• Used primarily to culture dermatophytes – molds that cause skin, hair and nail infections
• Should not be used as a primary plating medium from all sites, for cycloheximide prevents
growth of some genera of pathogenic fungi:
• Trichosporon species, Candida tropicalis and Cryptococcus neoformans and C. gattii
• Brain heart infusion (BHI) medium /with or without blood
• Used for the primary recovery of fungi
• Inhibition of bacterial growth with addition of chloramphenicol and gentamicin
• Sheep’s blood can be added to the medium to nurture for fastidious systemic fungi such as
Histoplasma capsulatum
6

Mycology Laboratory Safety Precautions
• BSL-2 level safety precautions
• BSL-2 biosafety cabinet with HEPA filtration required for working
with mold cultures
• Yeast procedures can be performed on bench-top
• PPE (personal protective equipment) include lab coat and
gloves
• Biohazardous waste must be autoclaved
• Agar plates must be sealed with oxygen permeable seal
after inoculation to protect those working in the
laboratory from spore formation during culture
incubation.
7

•Using Biosafety Level 2 precautions, inoculate specimen onto selected media
and streak for isolation
•Seal plates with oxygen permeable tape prior to incubation
•Tubes or vials can also be used for culture in place of agar plates, but it is
more difficult to visualize fungal growth and perform testing
•Incubate media at 30˚C for 4 weeks (extended time necessary for the
detection of slower growing systemic fungi), screen periodically
•If growth occurs - perform appropriate identification tests
Processing of Fungal
Cultures
8

Yeast Identification Methods
• Biochemical reactions have been used for decades to identify yeast.
Recent taxonomic updates and new emerging yeast pathogens have
challenged the accuracy of biochemical identification methods.
• Newer methods with improved accuracy of yeast identification include:
(1) MALDI-tof
(2) RNA sequencing – particularly helpful for rare and difficult to
identify yeast species that are not in the MALDI-tof library. RNA
sequencing (RNA-Seq) analyzes the sequence of the yeast’s
ribosomal RNA (rRNA), which contain unique sequences that
allow for species-level identification of different yeast strains
9

MALDI-tof
(Matrix Assisted Laser Desorption/Ionization Time of Flight)
Yeast colony is smeared onto the target plate and a matrix solution is pipetted
onto smear spot and allowed to dry. Target plate is placed in instrument. Laser
beam superheats each smear spot separately, and converts the yeast into
charged protein particles, the particles based on mass and charge travel up a
vacuum chamber to a detection plate to form identification peaks that identify
yeast to genus and species. Identifies most yeast infecting humans.
10

• Lactophenol Cotton Blue [LCB] adhesive tape preparation used for
decades to presumptively identify molds using microscopic morphology
• LCB mounting medium consists of phenol, lactic acid, glycerol and
aniline cotton blue dye.
• Clear adhesive tape is used to touch the colony, picking up fungal
hyphae/conidia, then pressed into one drop of LCB on a microscope
slide. Observe under light microscope using 40X.
• Can be used for presumptive identification to genus for numerous
molds.
Newer/ more definitive
identification methods include:
(1) MALDI-tof – extraction method
prepares mold for testing
(2) RNA sequencing techniques
Mold Identification Methods
11

Gram stain
•Most yeast species stain well with Gram stain, staining Gram positive (blue)
•Examination for budding cells can assist with confirmation that it is a yeast and not a
staining artifact
•Pseudo-hyphae Gram stain well, they are formed by yeast cells elongating with incomplete
budding, yeast cells remain attached to pseudo-hyphae
•Mold mycelia may be difficult to visualize on Gram stain slides
pseudohyphae
Mold mycelia
Yeast
Pseudo-hyphae
Direct Visual Examination of
Specimens
X100 oil immersion
12
Budding yeast cells

•Used to detect yeast, pseudo-hyphae, and hyphae most usually in skin,
hair, and nail specimens
•KOH dissolves the keratin in cellular material and frees hyphae and yeast
cells so they can be better visualized on the slide preparation
•Moderate sensitivity and specificity particularly for mold, the colorless
hyphae can be difficult to visualize and can be confused with cell borders
•Used less as better techniques evolved
Potassium Hydroxide Prep (KOH)
X40
Light microscopy
13

• Yeast, pseudo-hyphae, and
mycelial fungi bind with the
Calcofluor White stain reagent and
fluoresce a bluish color
• Stain is read using a fluorescence
microscope (40X)
• More sensitive and specific method
than a KOH preparation for
visualization of yeast, pseudo-
hyphae and hyphae
Calcofluor White
Stain
14

One drop of black ink is placed into one drop of CSF on a
microscope slide, cover with a coverslip and examine
using light microscopy (40X) (can also be used to examine
other body fluids)
“Negative” staining technique – stains the background only
for the ink is unable to penetrate the yeast cell or capsule.
The polysaccharide capsule is produced by both
Cryptococcus neoformans and C. gattii.
Less sensitive method than the Cryptococcal antigen test.
Cryptococcal antigen is considered the superior test for
diagnosis of Cryptococcal meningitis.
India Ink – Cryptococcus detection in CSF
15

Molecular and Sequencing for Direct Detection
• Direct detection of fungal microbial DNA from patient specimens:
• DNA is isolated from specimen and amplified by PCR using a battery of broad-range
primers for yeast and mold organisms
• Amplified products are sequenced for organism identification
• Detection and identification of specific targeted pathogens:
• DNA is isolated from a cultured organism or direct patient specimen and amplified by PCR
using primers developed specifically to search out a particular yeast or mold.
• Liquid biopsy to detect a large menu of infectious agents from plasma.
• This technique detects microbial cell–free DNA circulating in the bloodstream from more
than 1000 pathogens that cause infections, including yeast and mold species.
Analysis of complex genomic data arrives at identification of the potential pathogen.
• These tests are offered at reference laboratories.
16

Helpful for the description of cellularity in tissue
associated with fungal infection, morphologic
assessment of size, septations and branching
pattern of molds and the size and budding
pattern of yeast. Can also visualize structures
such as sulfur granules.
Hematoxylin and Eosin Stain
17
Sulfur Granule
Aseptate fungal hyphae
Granuloma
Examination of Fungi in Fixed tissue
Intracellular Yeast

Morphology of Molds in H & E Stains
Narrow, uniform Broad, irregular hyphae Pigmented hyphae/spores
hyaline septate hyphae folded appearance
Aspergillus Mucorales such as Dematiaceous fungus
Fusarium Mucor and Rhizopus Culture for definitive
Scedosporium Culture for definitive identification
Culture required for identification
definitive identification
18

Grocott’s Methenamine Silver Stain [GMS]
Yeast and hyphae stain grey to black and stains both living and dead fungi.
• Molds: Observe the width of the hyphae, the presence or absence of regular
septations, and the angle of branching
• Yeast: Observe the size and budding pattern and presence of pseudo-hyphae
• Later slides will discuss specific observations that assist in identification
19

PAS stain yeast and hyphae magenta. PAS is not specific to fungi, will
also stain structures containing carbohydrate macromolecules
(glycogen, glycoprotein, proteoglycans). PAS stains only living fungi.
Periodic Acid Schiff [PAS] Stain
20

Stains the polysaccharide capsule of Cryptococcus neoformans and
C. gatti a pink color and can be used to confirm identification.
Mucicarmine [Mucin] Stain
Fontana-Masson Silver Stain detects melanin, used in the rare situation
when capsule deficient cryptococcus organisms are encountered.
Non-encapsulated Cryptococcus stain brown.
21

Antifungal Drug Classification and Action
• Loss of cell membrane integrity:
• Polyenes: Amphotericin B and nystatin
• Action: Bind to ergosterol, a steroid-alcohol unique to Fungi to create pores in the fungal cell
membrane, leading to electrolyte leakage and cell death
• Azoles: Ketoconazole, miconazole, clotrimazole, itraconazole, isavuconazole, fluconazole,
voriconazole, and posaconazole
• Action: Inhibitors of the fungal enzyme lanosterol 14-alpha-demethylase, which is a rate-limiting
enzyme in the fungal biosynthetic pathway of ergosterol. This action destabilizes the fungal cell
membrane, causing cell content leakage and death.
• Loss of cell wall integrity:
• Echinocandins: Anidulafungin, caspofungin, and micafungin
• Action: Inhibit the fungal beta-(1,3)-D-glucan synthase, which is the enzyme responsible for
synthesizing beta-(1,3)-D-glucan, a key component of fungal cell walls. Losing this cell wall
component leads to osmotic instability and cell death.
• Susceptibility testing can be performed to assist in therapy selection.
22

DIMORPHIC FUNGI
Systemic pathogens with unique characteristics
24

Dimorphic Fungus?
• Depending on temperature and conditions of environment, one fungi
demonstrates two forms:
• (1) Mycelial form - Hyphae with conidia (spores)
• Free living form in nature and growth in laboratory at temperatures <=30˚C
• (2) Yeast or yeast like form
• Parasitic phase found in human tissue and growth in the laboratory at temperatures >= 35˚C
• Exposure to increased body temperature transforms the mycelial form acquired in nature to
the yeast form that infects tissue
Histoplasma capsulatum –
Mycelial form in nature and growth at <=30˚C
Histoplasma capsulatum – Pathogenic
Yeast form in tissue and growth at >=35˚C
25

Dimorphic Fungi
• Histoplasma capsulatum and H. duboisii
• Blastomyces dermatitidis
• Coccidioides immitis and C. posadasii
• Paracoccidioides brasiliensis
• Sporothrix schenckii complex
• Talaromyces (Penicillium) marneffei
26

Epidemiology: Worldwide distribution,
in the USA primary endemic region is Ohio,
Missouri, and Mississippi River valleys, but global
warming has led to an expansion of the region.
Environmental sources: Aerosolized bat guano is rich in H. capsulatum
conidia, so infections occur in spelunkers (cave explorers). Conidia found in
many species of bird droppings, including chickens, making farmers at risk.
27

Histoplasmosis
• Up to 90% of infections are asymptomatic or subclinical
• @ 10% acute pulmonary, chronic pulmonary or disseminated
with primary sites: bone marrow, heart, CNS, gastrointestinal,
skin & genitourinary
• Mucocutaneous lesions a unique dissemination site
• Who is most likely to have disseminated Histoplasmosis?
• HIV/AIDS / both primary and reactivation disease
• Organ transplant / both primary and reactivation disease
• Taking medications such as corticosteroids or TNF-inhibitors
• Infants
• Adults aged 55 and older / both primary and reactivation disease
• Treatment for serious infections include : Itraconazole,
Fluconazole +/- Amphotericin B
28

Histoplasmosis Diagnosis
• Antigen detection in urine and/or serum
• Quantitative enzyme immunoassay
• Performed on random collected urine or serum specimen
• Most sensitive (>=85%) for the detection of disseminated or chronic
pulmonary Histoplasmosis cases, can detect early in infection
• Useful test in immune suppressed patients that do not produce a
detectable antibody response
• H. capsulatum antibody tests are available (immunodiffusion and
complement fixation.) Antibody usually appears in the second month of
infection so not as useful for acute diagnosis as antigen detection.
29

H. capsulatum Culture - Mold
• Infection acquired by inhaling microconidia found in
nature / capable of penetrating into the lung
• Requires 2-8 weeks to grow in culture at 30*C
• Colony is white to brown and cottony
• Microscopic appearance on LCB tape mount
• Slender septate hyphae
• Tuberculate macroconidia, large & round (8 – 16 µM)
• Microconidia (2 - 4µM) (infectious particle)
• MALDI-tof, DNA hybridization probe, or 16S rRNA
sequencing necessary to confirm identification
• Confirmation is required due to look alike fungi, such as
Chrysosporium and Sepedonium species, which are usually
considered to be saprophytic not disease producing.
30
Large Macroconidia have
tubercles (spike protrusions)

H. capsulatum Culture-Yeast
• Requires 4-8 weeks to grow in culture at 35*C
• Colony is white to beige with a creamy consistency
• Microscopic appearance
• 2-4 uM in size, oval to round, narrow-based budding
• Yeast is the form seen in human tissue
• Increase in temperature in human induces the inhaled conidia from soil to undergo
transformation to yeast in the host’s tissue
31

Histopathology H. capsulatum
•Granulomas produced in lung and other tissues can be either non-
caseating or caseating
•Primary infection usually in lung but can disseminate to organs of
the Reticuloendothelial System (RES) – high % of cases with
dissemination to bone marrow
32

Histoplasma capsulatum – Yeast in fixed tissue
•Small yeast 2-4 uM, minimal size variation, narrow neck bud, oval to round.
•Yeast are often intracellular within macrophages
•In H& E stain, yeast appear to be encapsulated due to staining artifact –
caused by membrane shrinkage away from the cell wall
•Stains well with a variety of pathology stains
•Possible confusion due to size with Nakaseomyces (Candida) glabrata,
Candida auris, pneumocystis, and Cryptococcus spp.
H & E
PAS
Gram Wright’s
33
Appears to be
encapsulated
GMS

Leishmania donovani
amastigote
Note: kinetoplast (mitochondrial
DNA) next to nucleus, L. donovani
does not stain with GMS
Toxoplasma gondii
tachyzoites - Oval to
crescent in shape, no capsule-
like clearing around tachyzoite
Beware of these Look-a-likes and rare occurrences!
34
in aortic valve tissue. Unlike in
other organs, Histoplasma can
produce variable size yeast cells
and hyphal forms on valve tissue in
cases of endocarditis.
This could be potentially confused
with other molds.

Case History: Histoplasmosis
38 yr old male, Newly diagnosed with HIV/AIDS
HIV Viral Load = 220,000 copies
CD4 count = 3
Anemia / Thrombocytopenia
Bilateral pulmonary infiltrates
Bone marrow aspiration procedure: Giemsa stain
and GMS stain of aspirate show small yeast cells
Culture performed / mold examined with LCB prep
35
Therapy: mild cases Itraconazole, serious or immune
suppressed add Amphotericin B.
Giemsa
stain
GMS stain
Lacto-phenol
cotton blue prep

Mold phase is identical in culture growth and
appearance on LCB prep to H. capsulatum.
Difference is the size of the yeast cell:
Note yeast cell of H. duboisii is 6-12 um,
which is 2-3X the size of H. capsulatum yeast.
Endemic area: Central, east and west Africa
Primary infection: cutaneous lesions with
occasional involvement of the bone
Unusual variant of Histoplasma –
Histoplasma duboisii
36

Blastomyces dermatitidis
• Epidemiology
• In the US, Ohio and Mississippi River valley and Great
Lakes region
• Infection is not associated with any animal or specific
activity
• Mold found in forested areas and riverbanks
• Infection from inhalation of airborne conidia
• Infects humans and dogs
• Primarily a pulmonary pathogen
• Small % of patients disseminate to skin and bone
• Well demarcated (raised border) skin lesions
• Dissemination in immune suppressed patients
37

Blastomyces Serology
• Blastomyces antibody testing from serum: testing methods available
for the detection of antibodies by enzyme immunoassay(EIA) and
immunodiffusion
• Serology testing is performed in conjunction with histology or
culture.
• Blastomyces antigen in urine: Diagnose and monitor B. dermatitidis
disease
• Urine specimen collected in sterile container
• Good test for immune suppressed with poor antibody production
38

Blastomyces dermatitidis - Mold
• Growth in 2-3 weeks @ 30 *C
• Fluffy white – buff colored mold
• Microscopic on LCB prep - pear shaped conidia at the end of
supporting hyphae. Looks like a lollipop
• Look-alike fungus – Chrysosporium species and possibly others
require confirmation of identification: MALDI-TOF, DNA hybridization
probe or 16s RNA sequencing
Blastomyces Chrysosporium
39

•Slow growing yeast taking up to 4 weeks to grow at 35*C
•Large yeast cell with size variation (8-15 um), round to slightly oval
•Unique “Broad Based Budding” pattern and thick/double contoured wall.
Blastomyces dermatitidis - Yeast
Broad based bud
Double contoured wall
40

Histopathology Blastomyces
dermatitidis
• Mixed neutrophilic and granulomatous inflammation is observed in tissue
• Stains well with H&E, PAS and GMS stains. May stain weakly positive on
mucicarmine and cause confusion with C. neoformans and C.gattii
• Classic Broad based budding yeast cell with thick wall
41
PAS stain
GMS Stain

Coccidioides immitis and C.
posadasii
• These two species share close genetic relatedness
• Found in different geographic regions, C. immitis mostly in
California region and both species found outside California
• Both species have the same infectious process, react the same in
serologic tests, and are morphologically identical
• Coccidioides endemic in SW USA (San Joaquin Valley), Mexico, and
South America, in areas known as the Sonoran life zone with warm
and desert sands, common in rodent burrows in the desert areas.
Global warming has led to an expanding endemic region.
• Infection is from inhalation of fungal particles (arthroconidia) found in
the sandy soil of the Sonoran life zone areas
42

Coccidioidomycosis
• 65% of patients have asymptomatic infection or flu-like symptoms
• 30% of patients have progressive pulmonary disease, may include
erythema nodosum (tender lumps under the skin), or may a coin lesion in
the lung
• Usually <5% will experience disseminated infections
• Infection of skin, bones, joints, lymph nodes, adrenal glands, and tropism to CNS
• Risk factors for severe or disseminated coccidioidomycosis include:
• Darker skinned individuals and those of Filipino descent
• HIV/AIDS
• Use of immunosuppressive medications
• Organ transplant
• Diabetes mellitus
• Pregnancy
43

Serologic diagnosis of Coccidioidomycosis
• Enzyme immunoassay (EIA): Sensitive and commonly used method for
diagnosing coccidioidomycosis, detects IgG and IgM antibodies
• Immunodiffusion (ID): detects IgM antibodies; positive early in the course
of infection,
• Not as specific as EIA detection of antibodies and can have “false” positive
results, must confirm infection with additional testing
• Complement Fixation (CF): detects IgG antibodies and allows for
assessment of disease severity.
44

Coccidioides Culture - Mold
• Growth in 2-3 days as a waxy colony at 35*C, becoming wooly in around 7–10
days
• Microscopically (40X) one observes areas with septate hyphae and thick-walled
alternating barrel shaped arthroconidia
• Barreled arthroconidia breakoff from hyphae and become the infectious particle in
nature and is inhaled into the lung to initiate infection
• Beware! Laboratory workers must cautiously handle in BSL-2 biosafety cabinet,
reported source of acquired infection
40X
2-3 days 7-10 days
45

Coccidioides Look-a-likes
• Malbranchea species is similar to C. immitis and C. posadasii under
the microscope using adhesive tape prep method
• Must confirm any fungi suspected to be Coccidioides spp. using
MALDI-TOF, molecular DNA hybridization probe or 16S rRNA
sequencing
Coccidioides Malbranchea
46

Histopathology Coccidioides
• No yeast cell produced in human tissue: Instead, the observed structure is
a thick-walled spherule variable in size (10 – 80 uM) with endospores.
• Spherules can be in all stages of development: fragmented to well formed
mature spherules with endospores (2-5 uM)
• Hyphae rarely observed, possibly in CNS infections
• Granulomatous inflammation with caseation
47
Position of two Coccidioides
spherules pushed together can
mimic the broad-based bud of
Blastomyces

Case History:
Coccidiodoimycosis
75 year old male
Right pleural effusion
Lung nodule
Past medical history: Gout,
hypertension, anemia, obesity
Fungal serology testing:
Histoplasma urine Ag: negative
Cryptococcal Ag: negative
Coccidioides IgM: Indeterminate
Coccidioides IgG: Weak Positive
Titer 1:4
Epidemiology: Lives on ranch in Simi
Valley, California
Biopsy of lung
nodule stain/ GMS
Culture of lung
nodule incubated
at 30°C
48

• Rhinosporidium seeberi (an aquatic parasite) also forms spherules
but they are larger than those of Coccidioides
• Rhinosporidium spherules can be > 80 uM in size compared to those of
Coccidioides <=30 uM
• R. seeberi cause oral or nasal mass lesions
Oral /nasal mass lesions
Organisms possibly confused with Coccidiodes in
tissue!
49
Prototheca wickerhamii
produce structures known
as morulas that mimic
spherules. Endospores
(2-20) are in a rosette
structure.

Paracoccidioides brasiliensis
• South American Blastomycosis – 80% of cases reported
from Brazil
• Most prevalent systemic fungal infection in Latin America
• Infection acquired from inhaling infectious particles from
soil and decaying vegetation or possibly trauma, leading
to skin infections.
• Disease presentation:
• Pneumonia
• Cutaneous lesions
• Disseminated infection can occur
• Extrapulmonary lesions on the face and oral mucosa are
common sites of dissemination
50

Paracoccidioides – Culture
• Cultures for mold are usually not performed due to slow growth
and nonspecific sporulation
• Yeast grows at 35-37˚C
• Slow growing – 3 weeks
• Large (10 – 30um), thick walled, with 2 or more tear drop shaped daughter
buds (2–10 um)
• Unique multiply budding yeast cell known as the Mariner’s wheel or
Pilot’s wheel yeast Careful! Do not confuse with tuberculate
macroconidia of mold phase of Histoplasma capsulatum.
51
Histoplasma capsulatum –
tuberculate macroconidia

Histopathology
Paracoccidioides brasiliensis
• Granulomatous non-necrotizing and
necrotizing or pyrogranulomatous response
• Mariner’s wheel yeast forms
GMS
GMS
52

Sporothrix schenckii complex
• Sporotrichosis
• Cutaneous inoculation from penetrating
injury or scrape with contaminated plant thorns
• Known as “Rose gardener’s disease”
• Usually begins as skin lesion with or without
ulceration and leads to a progressive
subcutaneous disease
• Lesions can progress with lymphocutaneous
spread and possible dissemination to bone and
other organs
• Pulmonary and CNS infections occur, but rarely
53

Sporothrix schenckii complex
culture
• Mold grows in 3-5 days at 30˚C in 3 -5 days, beige
colony eventually turns brown/black
• Microscopic: thin septate hyphae with conidia in
daisy wheel or rosette pattern
• Yeast grows in 7 days at 35˚C as oval to elongated
yeast cells, 2 x 5 µm, described as cigar bodies
54

Histopathology S. schenckii
• Pyogenic to granulomatous inflammation
• Yeast may not be observed in human tissue
• If seen, elongated, pleomorphic described as cigar shaped
• More common structure seen in human tissue is an Asteroid body also
known as Splendore-Hoeppli phenomenon (SHP)
• SHP are not unique to Sporotrichosis, also seen in infections with:
• Mucormycetes (Mucor, Rhizopus)
• Aspergillus
• Blastomyces
• Candida spp
55
Asteroid body Pleomorphic yeast cell

•Disease: Cutaneous bumpy lesions
(Talaromycosis) most common but pneumonia
or systemic infection can occur in immune
deficient or HIV/AIDS
•Endemic in the soil in SE Asia, southern
China and eastern India
•Mold grows in 2-3 days (<=30*C) as a green
mold with red diffusible pigment
•Mold on LCB prep looks like a typical
Penicillium species with branched
conidiophore and attached conidia
•Fixed tissue stain shows small yeast-like cells
that divide by transverse fission
Talaromyces (Penicillium) marneffei
56

57
Mold In Tissue
Summary of important molds appearance in tissue
Appearance of organism Appearance of pathology

SUBCUTANEOUS FUNGAL
INFECTIONS
Mycetoma
Chromoblastomycoses
Phaeohyphomycosis
Protothecosis
58

Mycetoma
• Chronic, granulomatous disease of the skin and subcutaneous tissue,
sometimes involves muscle, bone, and neighboring organs
• Occurs in hot temperate parts of the world
• Trauma to skin occurring in nature implants the fungi into the
subcutaneous tissue, farm workers are most at risk
• 3 criteria help to explain mycetoma:
• Swollen extremity with lesion progression
• Development of abscess and fistulas (sinus tracts)
• Sulfur granules drain from sinus tracts, observed in tissue histology slides to help
establish diagnosis
Two types of mycetoma:
1. Actinomycotic – infection with higher bacteria species
2. Eumycotic – infection with black molds
59
Draining sinus with
sulfur granule

Actinomycotic Mycetoma
• 98% of all mycetoma infections are actinomycotic
• Nocardia species most common infectious agent
• Sulfur granules are formed in tissue.
• The granules are composed of a matrix of the filamentous
bacteria. Observe the edge of stained granule to observe
the size and morphology consistent with Nocardia species
60

Edge of sulfur granule show thin filamentous
bacteria: How can you tell if infection is due to
Nocardia or Actinomyces?
Perform a PAF stain and culture:
•Nocardia species are modified (partial) acid fast
[PAF] stain positive and grow aerobically
•Actinomyces species are PAF stain negative and
grow best anaerobically
Careful! Sulfur granules created by infection
with Actinomyces species (an anaerobic
Gram-positive bacilli) can appear very similar
to ones formed by Nocardia species, but this
is a distinct infection known as Actinomycosis.
Sulfur granules in Tissue
Actinomyces
61
Nocardia

• Aerobic, filamentous, branching Gram positive bacilli.
• Nocardia can stain poorly with Gram stain and appear speckled
• Modified Kinyoun Acid-Fast Stain (PAF) stains red /Partial Acid-Fast
positive
Modified (Partial)
Kinyoun acid-fast stain
Gram stains
Nocardia species
62

• Besides Mycetoma, Nocardia spp can cause primary
Pulmonary and Brain infection in immune suppressed
• Grows in 3-5 days at 30 – 35*C on many agar types
including SABs and 5% Sheep’s blood agar
• Colony is dry/crumbly with a musty odor and can vary
in color, but usually white to coral
• There are 85 species of Nocardia with the members of
Nocardia asteroides complex being most common
• Identification: MALDI-TOF & 16s rRNA sequencing
• Therapy is species specific and best guided by
performing susceptibility testing
Nocardia species
63

Thermoactinomycetes
• Gram positive filamentous rods related to Nocardia species
• Thermoactinomyces and Saccaropolyspora species
• Flourish in areas of high humidity and high temperature (40*-60*C)
• Grow best in moist and hot agricultural environments, like stacks of hay
• Agents of the disease Farmer’s lung – hypersensitivity pneumonitis or extrinsic
allergic alveolitis, occupational disease of farmers
• Immunologically mediated inflammatory disease of the lung
• Inhalation exposure to the thermophilic actinomycetes
• Pathology: Loose, non-necrotizing granulomas
• Diagnosis
• Detailed environmental history
• Serology testing
64

Eumycotic Mycetoma
Infection with pigmented/black molds labeled dematiaceous molds
-Numerous species of black molds found in soil and soil debris
-Mycetoma infection from the implantation of the mold into the
subcutaneous tissue, usually a result of traumatic event in environment
- Cause only @ 2% of mycetoma-like infections
-Most common agents: Acremonium spp. and Scedosporium
apiospermum complex
Notice the thick
hyphae on the
edge of the granule
65
Dark grey black colony

Black molds /known as Dematiaceous
molds
•Black colored colony on top and reverse side
•Black due melanin or melanin like pigments in the cell
wall of the mold
•Commonly found in areas damaged by flooding
•Commonly found in the environment
•Difficult to identify based on morphology due to
different fungal genera sharing sporulation
characteristics, usually require MALDI-TOF or
sequencing for identification
66

Exophiala species
A few common Black Molds
Cladophialophora bantiana
Associated with dissemination /
can extend to brain
67
Wangiella dermatitidis
Phialophora
verrucosa
Fonsecaea species
Acremonium species

Chromoblastomycosis
• Three characteristics help in the diagnosis:
• Wart like lesions into subcutaneous tissue (scarred and nodular)
• Brown septate muriform bodies observed in fixed tissue slides
• Infection caused by black pigmented fungi (dematiaceous)
• Skin trauma implants fungi into tissue causing infection
In tissue: Naturally brown, septate
structures known as muriform bodies,
sclerotic bodies, or copper pennies.
These structure can break up into pieces
and extend into the surrounding
subcutaneous tissue spreading infection.
Most common agents: Fonsecaea,
Cladophialophora, and Phialophora spp
68

Prototheca wickerhamii (Protothecosis)
• Algae without chlorophyll (not a fungal organism)
• Related to penetrating injury occurring in nature
• Causes nodular skin lesions and olecranon bursa infection
• Most common in patients with suppressed immune system
• Compare morula of Protothecosis to copper penny of Chromoblastomycoses
• Morula of Protothecosis is NOT naturally brown in color and has no regular septations
Prototheca
morula
Copper penny
69
Chromoblastomycoses

Phaeohyphomycosis
Traumatic implantation of black pigmented fungi into subcutaneous tissue
• Infections usually nodular lesions or cysts on the skin, controlled by an
intact immune response but may disseminate in the immune suppressed
• Disseminate to lung, or primary nasal sinus infection with extension to brain
• In fixed tissue, dark brown colored swollen hyphae and yeast-like cells
• Most common agents: Alternaria, Curvularia, Exophiala and Phialophora
70

Alternaria species
•Commonly found in nature in soil and plants
•Causative agent of phaeohyphomycosis (skin and subcutaneous
tissue), sinus, and nail in the normal host
•Emerging pathogen for invasive disease in immune suppressed
•Grows in 3 – 5 days at 30*culture on a variety of fungal media
•Black colony producing multicellular chaining conidia
71
Scotch tape exam

Curvularia lunata
Center cell is the largest
• Found on soil and plants in tropical and subtropical areas
• Phaeohyphomycoses, erythematous skin lesions (Eumycotic
mycetoma), sinusitis, and invasive infections including endocarditis.
• Emerging pathogen in the normal and immune suppressed
• Grows in 3-5 days at 30*C in culture on variety of fungal media
72
Scotch Tape Exam

Exserohilum rostrum
• Causative fungus in an outbreak from a compound pharmaceutical [steroid]
product contaminated during manufacturing with environmental dust
containing Exserohilum
• Contaminated commercial product injected into lumbar spine and knee joints
for pain management and led to serious and fatal infections:
• Meningitis
• Spinal abscess
• Synovial infections
73

Scedosporium apiospermum (Pseudallescheria boydii complex)
•Found in soil and environment
•Eumycotic mycetoma
•Pulmonary pathogen from spore inhalation with possible dissemination in immune suppressed
Can invade vessels, particularly in the lung, and lead to infarcts
•Cat fur-like gray colony with growth in 3 – 5 days at 30 *C on variety of fungal media
•Microscopic appearance: Lollipop-like spore production branching from septate hyphae
•Difficult to distinguish from Aspergillus in fixed tissue stains – both can branch at 45* angle
•Unique fact: Intrinsically resistant to Amphotericin B
74
PAS stain of fungal
hyphae
LCB preparation

IMPORTANT YEAST CAUSING
HUMAN INFECTION
Candida species
Cryptococcus neoformans and C. gattii
Trichosporon species
75

Candida species
• Greater than 200 species but @ 10 species common in humans
• Found as normal flora in GI, GU, and skin
• Opportunistic involving skin, mucous membranes and invasive sites
• C. albicans is the most common yeast causing human disease (@60%)
• Implicated in both superficial and systemic disease
• Nail, ear, endocarditis, urinary tract and bloodstream
• Risk factors include age of >=65 years, immunosuppression, leukocytosis,
intensive care unit stays, or presence of intravascular or urinary catheters.
76
Thrush

Candida species
• Growth in 24 – 48 hours at 30-35*C
• Growth on many agars SABS, IMA, Sheep’s Blood agar
• Bacteria-like colony – pasty white
• Oval shaped @ 7-8 um in size** with narrow neck bud
• Most species form pseudohyphae in human tissue. Not
true hyphae for the yeast do not detach, the yeast cells elongate to form
hyphae
• However, C. auris and Nakaseomyces (Candida) glabrata are two species
that do not produce pseudohyphae in tissue,** also small, 2-4µm in
size**
• Identify using biochemicals, MALDI-TOF, or 16 sRNA sequencing
pseudohyphae
77

C. albicans ID –
the old way
• Germ tube formation
• Incubate yeast in serum for 3-4 hrs at 35 ˚C
• Look for an extension from yeast cell (germ tube)
• Note: C albicans, C dubliniensis, and C stellatoidea can form
germ tubes and if incubation is extended >4 hrs – C.
tropicalis produces germ tubes
• Chlamydospore formation
• Growth on cornmeal agar at 48 hrs
• Form rudimentary spore (chlamydospore) structures unique to
C. albicans
chlamydospore
78
Starry appearing
Colony on BAP

Candida auris
• Nosocomial colonization and infection in critically ill patients with lines and tubes in
place or who recently spent time in skilled nursing or long-term care facilities
• Additional risk factors include those risk factors for other types of Candida infections: recent
surgery, diabetes, broad-spectrum antibiotic and antifungal use.
• Infections in patients of all ages, preterm infants to elderly with mortality 30 – 60%
• Very transmissible in healthcare settings through contact with contaminated
environmental surfaces or equipment, and from person to person
• Patients and healthcare workers can be colonized on skin, particularly axilla and groin, and
can persist for months
• Patient surveillance screening by PCR (most sensitive) or culture using
chromogenic media
• Intrinsic resistance to Fluconazole and Amphotericin B and some clades can also
be resistant to the enchinocandin class of medications (Micafungin)

80
Candida species Susceptibility to Azoles
(Fluconazole)
Unique features
C. albicans Susceptible Most common yeast / @ 60%
C. parapsilosis Susceptible Can cause invasive infection in
neonates and pediatric pateints
Nakaseomyces (Candida) glabrata 50% resistant Small yeast (2-4um) no
pseudohyphae produced
C. tropicalis 50-75% resistant
Pitchia
kudriavzevi (Candida krusei)
100% resistant
C. auris
C. lusitaniae
100% resistant
Susceptible
Emerging nosocomial pathogen,
Amphotericin B resistant,
Small yeast (2-4um) no
pseudohyphae produced
Resistant to Amphotericin B
Most common Candida spp in human infection and unique features

Candida Chrome agar plus for the preliminary
identification of Candida species:
When grown on this agar type, a unique color is formed
for six Candida species due to action on chromogenic
substrates contained in the medium.
Susceptibility testing of yeast is much like
bacteria, using broth dilution and Etest methods.
81
ChromAgar
C. auris

Candida Histopathology
• Pyogenic to granulomatous inflammation produced in tissue
• Most Candida species produce oval yeast cells (@ 8 um) with narrow
neck buds and possibly pseudohyphae in tissue.
• N. glabrata and C. auris form smaller yeast cells (2-4 um) and no
pseudohyphae is formed. Could be confused with H. capsulatum.
N. glabrata or C. auris GMS stain
Candida species not N. glabrata or C. auris
82

Cryptococcus
neoformans
• In nature, a 2um non-encapsulated yeast cell
• Found in pigeon droppings and soil contaminated with droppings
• C neoformans is enriched by the nitrogen, humidity and warmth of the heaped
droppings
• Small, non-encapsulated yeast cells are inhaled – travel through the
pulmonary system with hematogenous spread to brain and meninges
(neurotropic)
• Infects compromised hosts but as of late increasing reports in the more
normal host – one of the major AIDS defining infections, and less
common in corticosteroid use and chemotherapy.
83

Cryptococcus gattii – close relative of C. neoformans
• First Isolated from forested areas of the Pacific Northwest (British Columbia,
Washington, Oregon, and California)
• Associated with soil debris, decaying tree bark, and eucalyptus trees
• Infection of both normal and immune suppressed hosts
• Primarily a pulmonary disease,
• Tumor-like mass known as a cryptococcoma may form in lung or CNS
• Culture, biochemical & staining characteristics identical to C. neoformans
• One defining biochemical reaction for C. gattii
L Canavanine glycine brom-thymol blue medium
C. gatti = tuns blue C. neoformans = colorless
MALDI-TOF and 16sRNA sequencing will correctly ID both species
84

Cryptococcus neoformans
and C. gattii
• Variable sized yeast cells (2 – 20 um)
• Virulence factor: Polysaccharide capsule
• Polysaccharide capsule provides basis of diagnostic testing:
• India ink exam- CSF placed in one drop of black ink. It is a
negative staining method , staining the background but not the
polysaccharide capsule or yeast cell. Sensitivity >=50%
• Cryptococcal antigen test – Titer of capsular polysaccharide
leached off yeast cell and detected in CSF and serum
• Most sensitive test for diagnosis of meningitis with sensitivity
>=99%, can also follow patient recovery with decreasing titer of
polysaccharide present in the sample
85
Positive India Ink

Grows on SABS and IMA in
1-3 days at 30 – 35*C culture
Mucoid colonies due to production of
polysaccharide capsule
C. neoformans & C. gattii are the only yeast
that form brown colonies on Birdseed agar
due to phenol oxidase activity
Gram stain /
stains as blue blobs due
to the poor penetration
of gentian violet due to
the polysaccharide
capsule
Cryptococcus
species have a
positive urease
enzyme reaction
Cryptococcus neoformans and C. gattii
Negative
86
Positive

Cryptococcus in tissue
87
with visible clearing around
yeast cells due to capsule
Mucicarmine stains the
capsular polysaccharide of
and C. gattii.
GMS stain –Notice the yeast are
variable in size, narrow neck
budding
Fontana-Mason
stain/ stains the cell
wall of Cryptococcus, used do identify
Cryptococcus without a capsule

Yeast and Yeast-like Organisms in Tissue
Small/medium Variable size Large
(2 - 7 um) (2 – 20 um) (8 – 15 um)
Oval Round Budding Budding
Yes No Coccidiodes No Yes
Histoplasma
N. glabrata C. neoformans/gatti
Candida spp. (narrow neck) Broad based bud
Malassezia thick double wall
Sporothrix schenckii
Talaromyces (Penicillium) marneffei Blastomyces dermatitidis
Pneumocystis jorovecii
88

Pneumocystis jiroveci
• Yeast like fungus / cannot be cultured
• Used to be named Pneumocystis carinii and
considered a protozoan parasite
• Pneumonia in the immunocompromised host,
particularly HIV/AIDS / alveoli become filled with the
organisms. Colonization can occur in the elderly
• Diagnosis:
• Staining of Bronchial lavage, lung biopsy tissue,
induced sputum using direct fluorescent antibody
(DFA) or GMS stain, and PCR methods
• Positive reaction in (1.3)-ß-d-glucan assay, but
numerous fungi are also positive
DFA
89
GMS

Pneumocystis jiroveci (yeast like fungus)
could be confused with Cryptococcus –
Careful! Central nuclear staining in
pneumocystis, collapsed helmet forms.
C. neoformans/ C. gattii – no
nuclear staining
Cryptococcus vs.
Pneumocystis
Pneumocystis froth on H & E
90

Trichosporon spp
• T. asahii most common species
• Commonly inhabit soil, water, & vegetables
• Colonize skin, mouth and nails.
• Grows within 24-48 hrs in culture, becomes wrinkled over time
• Hyphae, pseudohyphae, arthroconidia, & rectangular yeast cells (3 x 8 uM)
• Urease enzyme positive
• Cause of superficial infection of the hair shaft named White Piedra, nail
(onychomycosis), and cutaneous infections
• Uncommon cause of fungemia and disseminated disease
• Most usually recovered from patients with corticosteroid use, solid tumors, HIV/AIDS, and
intravascular devices, including catheters and prosthetic heart valves
• Difficult to treat, elevated MICs to most anti-fungal drugs including echinocandins.
• Mortality 50 – 80%
91

CUTANEOUS AND
SUPERFICIAL MYCOSES
Malassezia furfur
Dermatophytes that cause Dermatophytoses
Microsporum species
Trichophyton species
Epidermophyton floccosum
92

Malassezia furfur
• Most superficial of the dermatomycoses
• M. furfur can be found on normal adult human skin and able to
colonize neonates at a very young age
• More common on oily skin or with high use of skin oils
• Most common in warm and humid climates
• Diseases:
• Skin infection is most common: Pityriasis versicolor / Tinea versicolor
• Macules, papules, patches, plaques on chest back and shoulders with either hypo
or hyper pigmentation – does not invade into deeper tissues
• Fungemia: caused by tunneling into bloodstream with IV catheter lipid
feeding lines (parenteral nutrition/hyperalimentation) in infants and adults
93

Malassezia furfur
• Lipophilic yeast – require long chain fatty acids for growth
• This can be provided on media with a film of olive oil
• Small budding yeast-like cells 2 – 4 µm with collarette (appears like
necklace at junction of mother and daughter cell)
• Skin and tissue preparations described as “Spaghetti and Meatballs”
due to yeast and short hyphal fragments.
94
Spaghetti and meatballs with
basket weave morphology in the
keratin layer
Spaghetti and meatballs

Dermatophytes – Ringworm infections
• Hair, skin and nail infections
• 3 genera of fungi:
• Microsporum species (many)
• Epidermophyton floccosum
• Trichophyton species (many)
• Disease described by area of the body infected:
• For example: tinea capitis (head), tinea pedis (foot)
• Usually, a clinical diagnosis but easily grown with
Mycosel being agar of choice. Maldi-tof capable of
genus/species identification
• KOH or Calcofluor white prep can be used to visualize
fungal hyphae from skin scrapings
95
Calcofluor white prep
Round “ringworm” lesions

Microsporum canis
•Skin or scalp ringworm infection acquired from an infected dog or cat
•Colony grows in 5-7 days at 30* C on Mycosel or other fungal media as a
white mold with yellow pigment on reverse side of colony
•Thick-walled macroconidia, tuberculate [tiny spiny projections]
•Rare microconidia (small spores)
96

Nannizia (Microsporum) gypseum
•Mostly scalp or skin infection from exposure to contaminated soil
•Sandy colored colony that grows in 5-7 days, 30*C on Mycosel or other
fungal media
•Large macroconidia are produced, no microconidia produced
97

Trichophyton rubrum
•Commonly infects skin and nails after
exposure to fomites, such as wet towels
•Grows in 5 – 7 days at 30*C. on
Mycosel and other fungal media.
•White mold, the back side of the colony
produces an intense red colored pigment
that diffuses into the medium
•Pencil shaped macroconidia and
abundant tear shaped micro-conidia
most common Can appear like “birds on
a wire” morphology along hyphae. Spiral
hyphae also present.
Red diffusible pigment
98

Trichophyton tonsurans
•White colony, grows in 5-7 days at
30* C on Mycosel and other fungal
media
•Light yellow on reverse side or agar
•No macroconidia
•Only microconidia with rare
ballooning microconidia which is the
identification structure
•Primary cause of epidemic scalp
ringworm in children, nails &skin
99

Epidermophyton floccosum
• Infections in skin and
nails
• Yellow-khaki green
colored colony that
grows in 5-7 days at
30*C on Mycosel and
other fungal media
• Beaver tail shaped large
macroconidia with no
microconidia produced
100

Opportunistic Hyaline Molds
Two groups based on hyphae
structure:
(1) Narrow uniform septate hyphae
(2) Broad non-septate ribbon like-
hyphae
Infections usually in the immune
suppressed host or special
circumstances
101

Hyaline molds with narrow septate
hyphae
• Hyaline – no color to the hyphae
• Regular septations along hyphae
• Grow on a variety of agar media
in @ 3-5 days at 30˚C or 35°C
• Preliminary identification based on growth rate,
color, texture of colony, and microscopic structures observed
on LCB prep
• Definitive identification: Using MALDI-TOF or 16sRNA sequencing
102

Serology Tests Supportive not Conclusive for
Diagnosis
Aspergillus Galactomannan Enzyme Immunoassay
• Detects circulating Aspergillus antigen in serum and/or bronchial lavage,
• Most diagnostic in advanced disseminated Aspergillus infections
• Moderate sensitivity and specificity (PPV 68%/ NPV 96%)
• False positive reactions (10%) occur with patients receiving Piperacillin/Tazobactam
therapy, infection with H. capsulatum, and rice and pasta ingestion
(1,3)–Beta-D-glucan assay
Polysaccharide found in the cell wall of many fungi
• Detects1,3 Beta-D glucan in serum or BAL with the following pathogens: Candida spp.,
Acremonium, Aspergillus spp., Coccidioides spp, Fusarium spp., Histoplasma capsulatum,
Trichosporon spp, Sporothrix schenckii complex, Saccharomyces cerevisiae, and
Pneumocystis jiroveci. Does not detect Cryptococcus or Zygomycetes
• High values more meaningful, false positive reaction can occur from colonization with fungi
103

Aspergillus species
• Ubiquitous in nature with @ 200 species
• Hyaline hyphae produced with regular septations,
occasionally irregular bulbous forms along hyphae
• Respiratory tract infection most common
• Can cause a fungus ball in the nasal sinus
• Can disseminate to other organs in immune suppressed
• Neutropenia can predispose to infection
Septations
104
Fungus ball

Aspergillus fumigatus complex
•Ubiquitous airborne fungus in nature, infection starts
by inhaling conidia
•Most common Aspergillus species causing pulmonary
infection in immune suppressed:
Aspergilloma, pulmonary nodules with cavitary formation
•Allergic bronchopulmonary syndrome in cystic fibrosis
and asthma, accompanied by elevation in both
eosinophil count and IgE
•Blue/Green colored mold, grayish with age
•Grows in 3 – 5 days at 30*C on multiple fungal media
•Phialides produce colorless conidia on upper third of
the swollen vesicle
106

Aspergillus flavus
•Normally isolated from cereals, grains,
legumes
•Same disease potential as A. fumigatus
but far less common, pulmonary infection
with possible dissemination, sinusitis and
nail infections
•Brownish green –yellow mold growing in
3-5 days at 30*C in culture
•Sometimes can visualize the Green/yellow
hue on conidia produced on phialides that
surround the swollen vesicle
107

Aspergillus niger complex
•Contaminant in fruits and vegetables, also
found in soil and decaying vegetation
•Chronic otitis media, occasional
aspergilloma and dissemination in
immunosuppressed
•Black colony with white border – visible
black fruiting heads grows in 2-5 days at
30*C on many types of fungal media
•Dark grey to black conidia produced,
supported by phialides that surround the
vesicle
108

Aspergillus terreus complex
•Isolated from soil and decaying vegetation
•Infection primarily in the immune
compromised host
•Sandy colored mold grows in 3-5 days at 30*C
on a variety of fungal media
•Colorless conidia supported by phialides,
headed upward
•Aleurioconidia produced (asexual spore
produced on the hyphae) are unique to this
species
•Note: Intrinsic resistance to Amphotericin B
109

Histopathology Aspergillus
Fruiting head seldom seen in fixed tissue
usually dichotomous (continuous) branching septate hyphae with
some pieces branching at a 45* angle, occasional bulbous hyphal
pieces. Invade vessels and unique calcium oxylate crystals.
Look a like fungus in fixed tissue
slides: Scedosporium species
Growth in culture will
differentiate the two fungi.
110
Invasion of vessels
Calcium oxylate
crystals

Neocosmospora (Fusarium) solani
•Common in nature, grows on plants
•White to fuchsia colored mold grows in 3-5 days 30*C on fungal
media. It is inhibited by cycloheximide (Mycosel media).
•Produces septate hyaline hyphae, banana boat shaped conidia
•Disease normal host: keratitis, nail and skin infections
•Disease immune compromised: sinusitis, pneumonia, deep
cutaneous lesions, and fungemia (unlike most fungi can be isolated
from blood culture)
•Random septate hyphae in fixed tissue
•Confirm identification with culture and identification
Banana boat shaped spore
111
Gram stain of positive
Blood culture
GMS stain tissue

Scopulariopsis species
•Found in soil and grows on plant
•Infections: Nail infections in normal host and skin, sinusitis,
pulmonary, brain abscess in immune suppressed host
•White to sandy colored mold growing in 3-5 days at 30*C on a
variety of fungal media
•Round, spiny conidia that chain
•Very resistant to antifungal agents including amphotericin B
112

Penicillium species – > 300 species
•Common in air, frequent laboratory contaminate
•Cause of bread mold
•Uncommon cause of human disease
•Can appear as a culture contaminate
•Blue/green colony grows in 3-5 days 30*C on a
variety of fungal media
•Hyphae with branched phialides that produce chains
of conidia / appears like brush or bony hand
113

The Mucorales
Mucor, Rhizopus and
related
molds
Fungi in the Order Mucorales
•Broad, Hyaline hyphae without septation
•Spores produced in a sack-like structure
known as sporangium
114

Mucorales
• Found in soil, rotten fruit and vegetables
• Unique infection: Acute rhinocerebral mucormycosis
• Occurs in diabetics due to the elevated glucose enriching growth of fungus,
usually caused by Mucor or Rhizopus species
• Infection starts in single sided nasal sinusitis then spreads to the same side orbit
of eye then invades the brain / high fatality rate
• Pulmonary infection in immune suppressed: most commonly
diabetics, corticosteroids and burn patients
• Molds grow within 24-48 hr producing coarse aerial hyphae
• Microscopically, broad, hyaline hyphae without septation seen with
spores produced within a sporangium (sack)
• Mince, do not grind tissue submitted for culture, the grinding will kill
the tubular non-septate hyphal structures
115

Rhizopus species Lichtheimia (Absidia) complex
Mucor species
No
rhizoids
Rhizoids
Species within the Mucorales
Distant rhizoid
116
Cunninghamella species
Mold grows within 24-48 hrs,
filling the plate with coarse
aerial hyphae

• Unique finding of 90˚ angle branching, with rare to no septations, ribbon
like hyphae, can invade vessels and cause infarcts and thrombi
• Faintly staining particularly on GMS stain
• Distinguishing hyphae characteristic of Mucorales, from other hyphae in
quite important, for Mucorales are more resistant than many fungi to the
azoles and echinocandin antifungal agents Invades vessels and
can cause infarcts
and thrombi
Mucorales in Tissue
117
90* angle branching

Mycology Update 2025 - Margie Morgan, PhD

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Mycology Update 2025 - Margie Morgan, PhD